An increasing body of evidence implicates somatic mutations as causally important in the induction of human cancers. These somatic mutations may accumulate in the genomes of previously normal cells, some of which may then demonstrate the phenotypes associated with malignant growth. Such oncogenic mutations may include a number of different types of alterations in DNA structure, including deletions, translocations and single nucleotide alterations. The latter, also known as point mutations, may frequently intervene in carcinogenesis, in that a variety of mutagenic chemicals induce such mutations. In addition, such mutations may occur spontaneously as a result of mistakes in DNA replication.
Point mutations have been directly implicated in the causation of 10-15% of human tumors. These tumors carry oncogenes of the ras gene family, which differ from their normal cellular counterpart protooncogenes by the presence of a point mutation at one of several sites in these genes. These mutations represent qualitative changes in the tumor cell genome which distinguish these cells from normal cells and provide a basis for diagnosis of the genetic origin of a tumor under study. Identification of the mutations that have created active oncogenes may provide important diagnostic and prognostic clues for tumor development. For example, a number of mutations have been found to alter the 12th codon of the ras oncogenes, causing replacement of a normally present glycine by any of a number of alternative amino acid residues. Such amino acid substitutions do not have equivalent effects; some substitutions (e.g., valine) create a potent transforming allele while others (e.g., proline) have only a limited effect on cellular phenotype. Thus, the identity of a particular nucleotide substitution may be a strong determinant of the behavior of the tumor cell (e.g., its rate of growth, invasiveness, etc.). As a result, DNA probes of oncogene mutations have promise as diagnostic reagents in clinical oncology.
Such probes are useful, however, only if a region of a gene of interest in which point mutations are likely to occur has been identified. Unless identification of such a region has been made, it is impractical to use oligonucleotide probes of limited size (e.g., 10-20 nucleotides long) to scan an entire gene, which might well be 30,000 or more base pairs long; for example, if a 15-nucleotide probe were used to scan the entire length of a gene 3,000-5,000 base pairs long, 3000-5000 separate probes would be required.
Thus, although DNA probes of oncogene mutations have potential as diagnostic tools, they cannot be used effectively unless a discrete region of mutation in the gene, which is causally related to activation of the gene's oncogenic function, has been identified. Without such localization of the region of mutational activation, use of DNA probes specific for point mutations is impractical.